Epstein-Barr virus (EBV) is an oncogenic human herpesvirus that has been linked to lymphoid and epithelial malignancies. The propagation and pathogenesis of EBV depends on the efficient activation of the viral lytic cycle. The primary initiator of the EBV lytic cycle is the immediate early protein Zta (also referred to as ZEBRA, BZLF1, and EB1). Zta is an atypical member of the basic-leuzine zipper (b-zip) family of DNA binding proteins with sequence similarity to the cellular C/EBP family, and functional homology with the Kaposi's Sarcoma Associated Herpesvirus (KSHV) K8 protein. Zta can bind to multiple sequence elements in the viral and cellular genome to stimulate transcription activity and origin-specific DNA replication. Zta can also pirate host factors necessary to create a permissive environment for viral replication. The ability of Zta to bind the viral origin of lytic cycle DNA replication (OriLyt) and nucleate a viral replisome suggests that is the EBV origin binding protein (OBP), and likely to share properties with the alpha and beta herpesvirus OBP UL9. New preliminary data indicates that Zta possesses single stranded DNA binding activity and can recognize a DNA hairpin that forms at the essential upstream element of OriLyt. Zta, like UL9, have a highly conserved and redox sensitive cysteine residue that is essential for replication function and single strand DNA binding. A redox sensitive cysteine is also essential for activation of C/EBP 2, a cellular factor that nucleates the KSHV lytic origin replisome. In this application, we propose to investigate the biochemistry, regulation, and genome biology of Zta in the reactivation of EBV lytic replication. We will test the overarching hypothesis that Zta shares biochemical and functional properties with other herpesvirus OBPs and that these common properties are essential for regulation and initiation of lytic replication. [unreadable] [unreadable] [unreadable]